CN104189009A - Small intestinal submucosa-based vascularization promoting thermosensitive material and preparation method thereof - Google Patents
Small intestinal submucosa-based vascularization promoting thermosensitive material and preparation method thereof Download PDFInfo
- Publication number
- CN104189009A CN104189009A CN201410440422.0A CN201410440422A CN104189009A CN 104189009 A CN104189009 A CN 104189009A CN 201410440422 A CN201410440422 A CN 201410440422A CN 104189009 A CN104189009 A CN 104189009A
- Authority
- CN
- China
- Prior art keywords
- small intestine
- sis
- submucous layer
- gel matrix
- matrix material
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Abstract
The invention provides a small intestinal submucosa-based vascularization promoting thermosensitive material. The small intestinal submucosa-based vascularization promoting thermosensitive material is prepared by packaging a small intestinal submucosa matrix and a dissolving solution, respectively, wherein the small intestinal submucosa matrix is obtained through the following steps: performing enzymatic hydrolysis, acid dissolution and freeze-drying on the small intestinal submucosa, and then crushing and sterilizing the small intestinal submucosa to obtain a sterile powdery or flocculent small intestinal submucosa matrix material; the dissolving solution consists of phosphate buffer containing NaOH or normal saline. According to the invention, the two matrixes of the small intestinal submucosa-based vascularization promoting thermosensitive material both can be preserved for a long time and are mixed in an appropriate ratio before in use; the thermosensitive material has excellent injectability and is capable of forming a stable gel at 37 DEG C without cross-linking polymerization; the thermosensitive material is proven to have the effect of promoting vascularization by both experiments in vitro and vivo.
Description
Technical field
The present invention relates to temperature sensing material of the promotion vascularization based on submucous layer of small intestine and preparation method thereof.
Background technology
Hydrogel (Hydrogel) is the gel taking water as disperse medium, there is the water content of the normal structure of approaching and good biocompatibility, physicochemical property is controlled, there is loose structure, its three-dimensional porous structure is very similar to extracellular matrix, is conducive to growth and proliferation of cell differentiation, is also conducive to the maintenance of bioactie agent activity, can Wicresoft implant, in organizational project, there is wide application prospect.Temperature-sensitive hydrogel can change gel state into from liquid state in certain temperature range, do not need other catalyst and violent reaction condition, can fill well various erose tissue defects, cell or active substance can be evenly distributed in to defect, the secretion of Promote cell's growth, extracellular matrix and the self-regeneration of tissue have outstanding advantage in organizational project and regenerative medicine application.Hydrogel can be by synthetic material (as PCL, PEG, PVA and PNIPAAm etc.) and natural material (as chitosan, collagen protein, hyaluronic acid and de-cell extracellular matrix etc.) preparation.Compared with the gel rubber material of synthetic, hydrogel prepared by natural material has better biocompatibility and biodegradability.
(the small intestinal submucosa of intestinal mucosa lower floor, SIS), be n cell epimatrix, thick about 0.8mm, collagen, Dan Baiduotang proteoglycan PG, glycosamine and the glycoprotein etc. that include complex array, can transmit molecule and cell information effectively.SIS has following characteristics: (1) SIS non-immunogenicity, do not cause immunological rejection for transplanting, SIS exceed 1000 kinds across kind of cross transplantation experiment in all show as non-immunogenicity; (2) SIS has antimicrobial acivity, can reduce infection; (3) SIS has good bio-mechanical property, and the SIS tensile strength of lyophilizing weakens, and by its rehydration 5 minutes, can reach stable mechanical state, is 1/7～1/14 of tendon intensity; (4) SIS has good biocompatibility, can promote various kinds of cell sticking, growing and breaking up on material, fast degradation in animal body; (5) SIS is mainly made up of I, III fiber type collagen protein, contain multiple somatomedin as basic fibroblast growth factor (bFGF), transforming growth factor-β (TGF-β), vascular endothelial cell growth factor (VEGF) and sulphuric acid proteoglycan, fibronectin (fibronectin, FN) etc., even if the SIS through preparation process processing still contains these somatomedin, there is the ability that promotes revascularization, tissue growth; (6) SIS has the ability of the tissue regeneration of site specific, and rapidly inducing cell infiltrates, and stimulates growing into and breaking up of angiogenesis and host cell, the regenerating tissues of generation on 26S Proteasome Structure and Function all to former similar in a organized way; (7) SIS convenient sources, is easy to preparation, and therefore, SIS is for tissue repair research, reconstruction peritoneum, bladder, tendon, blood vessel and cerebral dura mater, and for body surface reparation etc., and existing clinical practice.For further expanding the clinical practice of SIS material, SIS can be prepared as to thermosensitive hydrogel.Kang etc. are by after SIS acid-soluble, neutralization in vitro, lyophilizing, be mixed into suspension with PBS before use, after being injected in vivo, can form gel [In vivo release of bovine serum albumin from an injectable small intestinal submucosa gel.Kang, K.N.; Kim, D.Y.; Yoon, S.M., et al.Int J Pharm, 2011,32 (16): 3969-3976], but fail in vitro to form thermosensitive hydrogel.Hurst etc. preparation SIS gel, its method mainly comprises: enzymolysis, acid-soluble, 10mM HCl dialysis etc., sterilizing adopts chloroform dialysis sterilizing, then with aseptic NaOH neutralization, and acquisition gel [Hurst, R.E.; Hauser, P.J.; Kyker, K.D., et al.Suppression and Activation of the Malignant Phenotype by Extracellular Matrix in Xenograft Models of Bladder Cancer:A Model for Tumor Cell " Dormancy " .PLoS One, 2013,8 (5): e6418], the method is very loaded down with trivial details, use Qian Xu sterile working add aseptic NaOH with in close HCl; And the object of its dialysis is to make HCl in solution to reach 10mM, consuming time longer; The gel that in addition prepared by the method is unsuitable for long-term preservation.
To sum up, the SIS gel that at present prepared by method, in vivo can plastic, but is difficult to stablize in vitro formation gel, if be directly used in surface wound, can not make SIS effectively stop at wound surface, is unfavorable for the reparation to surface wound; And what conventional sterilizing methods adopted is the immersions such as 75% ethanol, chloroform, product form is liquid, is not suitable for a large amount of production [Unilted States Patent (10) Patent N0.:US 8,361,503 B2]; As in close rear preservation, even product at normal temperatures in time extend also can form colloidal state, lose its mobility, be unsuitable for long-term preservation; As the solution closing in preserving not, though can extend the holding time, need sterile working to add aseptic NaOH, and measure its pH value, complex operation, is unfavorable for application.
Summary of the invention
The object of the present invention is to provide and can form in vitro gel and there is temperature sensing material based on submucous layer of small intestine of Angiogensis and preparation method thereof.
Experimental study shows, during SIS Digestive system is direct and after lyophilizing, add again PBS solution to dissolve, all can not effectively form in vitro gel, but, by SIS host material and separately packaging of alkali liquor, when use, remix really plastic in vitro, inference thus, the order of neutralization reaction may be the key factor that affects its external plastic.
Based on above-mentioned inference, the invention provides the temperature sensing material based on submucous layer of small intestine, it is to prepare after submucous layer of small intestine gel matrix material, lysate are packed respectively, wherein, submucous layer of small intestine gel matrix material obtains by the following method:
Get submucous layer of small intestine, through enzymolysis, acid-soluble, lyophilization, beats powder after dry or shreds into cotton-shapedly, and sterilizing, obtains aseptic powdery or cotton-shaped submucous layer of small intestine gel matrix material;
Lysate: formed by the phosphate buffer that contains NaOH or normal saline.
In preliminary experiment process, find, adopt the separately mode of packaging really can promote the external plastic of product, still, not that any condition all can reach such effect: gel strength, lower than 2%, can not effectively form gel, higher than 4%, not unusual thickness when plastic, without syringeability.Therefore, the mass volume ratio of submucous layer of small intestine gel matrix material and lysate is defined as 2～4:100 (g/ml) by the present invention.
Wherein, in described lysate, the content of NaOH is as the criterion with adjusting submucous layer of small intestine gel matrix material pH=7.
Further, prepare the concrete operations of submucous layer of small intestine gel matrix material as follows:
(1) get dry submucous layer of small intestine, pulverize, gained SIS powder is for subsequent use;
(2) in HCl solution, add pepsin, treat that pepsin dissolves completely, obtains Digestive system;
(3) get SIS powder and Digestive system, after mixing, be placed on shaking table and rock, after Digestive system becomes clarification, take out and carry out lyophilization, beat powder after dry or shred into cotton-shapedly, sterilizing, obtains aseptic powdery or cotton-shaped submucous layer of small intestine gel matrix material.
Further, in step (1), after pulverizing, get the powder of 80 mesh sieves, be SIS powder.Under this particle diameter, be conducive to follow-up dissolving and digestion.
Further, in step (2), HCl pH value of solution=2～4; In Digestive system, pepsin concn is 1mg/ml.
Further, in step (3), the mass volume ratio of SIS powder and Digestive system is 1:100～2:100.(g/ml)。
In the present invention, submucous layer of small intestine gel matrix material adopts irradiation or oxirane to carry out sterilizing, and lysate adopts filtration sterilization.
The present invention also provides the preparation method of small intestinal submucosa gel, and it comprises following operating procedure:
(1) prepare submucous layer of small intestine gel matrix material:
A, get dry submucous layer of small intestine, pulverize, gained SIS powder is for subsequent use;
B, in HCl solution, add pepsin, treat that pepsin dissolves completely, obtains Digestive system;
C, get SIS powder and Digestive system, after mixing, be placed on shaking table and rock, after Digestive system becomes clarification, take out and carry out lyophilization, beat powder after dry or shred into cotton-shapedly, sterilizing, obtains aseptic powdery or cotton-shaped submucous layer of small intestine gel matrix material;
(2) after getting submucous layer of small intestine gel matrix material, lysate and packing respectively, obtain small intestinal submucosa temperature sensing material.
Temperature sensing material prepared by the present invention, two kinds of base materials all can be preserved for a long time, after before using, the two being mixed by proper proportion, there is good syringeability, 37 DEG C can form stable gel, do not need cross-linked polymeric, and this temperature sensing material experiment in vivo and vitro all confirms to have promotion vascularization.
Compared with the temperature sensing material of prior art:
(1) prior art can not be stablized in vitro and builds gel, and prior art fails to confirm to have retained growth factor activity; And material of the present invention can mix rear plastic in vitro, and can slow release somatomedin, inside and outside all can effectively promote vascularization, particularly can extend the effective acting time of SIS at surface wound, is conducive to the reparation to surface wound; Meanwhile, in some pre-stage test, can directly adopt external model to verify, the inconvenience of having avoided in vivo test to bring.
(2) in prior art, need to dialyse, just can make HCl in solution reach 10mM, thereby ensure plastic stability, it is consuming time longer.In this method, do not use dialysis, can form equally stable gel, it is more convenient to operate.
(3) what prior art sterilizing methods adopted is the immersions such as 75% ethanol, chloroform, and preparation process needs gnotobasis, is not suitable for a large amount of production; In the present invention, adopt irradiation or oxirane to carry out sterilizing to gel matrix material, buffer adopts filtration sterilization, is more suitable for a large amount of production.
(4) product that prepared by prior art is liquid solution or suspension, even if product extends and also can form colloidal state in time at normal temperatures, loses its mobility, can not preserve for a long time; And separately pack after sterilizing of the present invention, can preserve for a long time, when use, remix plastic, ensure the stability of product and the convenience of later stage use.
Brief description of the drawings
Fig. 1 SIS thermosensitive hydrogel plastic figure, wherein A, B are respectively concentration 2%, 3%SIS thermosensitive hydrogel
The scanning electron microscopic observation figure of Fig. 2 SIS thermosensitive hydrogel
Fig. 3 thermosensitive hydrogel percent hydrolysis testing result
Fig. 4 SIS gel growth factor slow release result
Fig. 5 SIS thermosensitive hydrogel promotes cell proliferation experiment result
Fig. 6 LIVE/DEAD cell fluorescence coloration result
Coloration result after Fig. 7 subcutaneous injection gel
Fig. 8 number of inflammatory cells order testing result
Microscopic examination result after Fig. 9 gel surface inoculation HUVEC
Figure 10 SIS thermosensitive hydrogel promotes rat aorta ring rudiment result
Figure 11 blood capillary testing result
The microscopic examination result of short Angiogenesis after injection in Figure 12 SIS thermosensitive hydrogel body
The impact of short Angiogenesis after injection in Figure 13 SIS thermosensitive hydrogel body
Detailed description of the invention
Embodiment 1 SIS temperature sensing material preparation
The SIS film of lyophilizing shreds, and through ball milling instrument ball milling 5min powdered under 25Hz, crosses 80 mesh sieves.Preparation 0.01N HCl (pH=2) solution, adds pepsin wherein, and in this pepsin digestion liquid, pepsin concn is 1mg/ml, on shaking table, places, and rocks to pepsin and dissolves completely.In Digestive system, add SIS powder, concentration is 1% (g/ml), places shaking table room temperature and rock after stirring, and observes SIS Digestive system and becomes after clarification thickness, is poured into culture dish, more than being placed into-40 DEG C of refrigerator pre-freeze 2h.SIS Digestive system good pre-freeze is put into freeze dryer, and lyophilization 24h is spongy.Spongy SIS digest is shredded into tiny cotton-shaped submucous layer of small intestine (SIS) gel matrix material.
In the present invention, adopt irradiation or oxirane to carry out sterilizing to gel matrix material, lysate adopts filtration sterilization.SIS gel matrix material and the phosphate buffer that contains NaOH (being lysate) are separated to packaging, obtain temperature sensing material of the present invention.
The SIS gel formula following (taking 1ml gel as example, every batch of SIS digest is as the criterion to test gained formula at every turn, and reference data is below provided) of preparation variable concentrations:
Table 1
When use, use the phosphate buffer that contains NaOH to mix with SIS gel matrix material, to pH approximately 7 left and right, be mixed with pregel suspension.After preparing, stir solid is fully dissolved, place 37 DEG C of incubators, within 20-30 minute, get final product plastic (seeing Fig. 1).
The present invention studies discovery, and gel strength, lower than 2%, can not effectively form gel, and higher than 4%, not unusual thickness when plastic, without syringeability.
In test, checking is found: during SIS Digestive system is direct and after lyophilizing, add PBS solution to dissolve, all can not form in vitro gel.
Illustrate beneficial effect of the present invention by test example below.
Test example 1
1, physics and chemistry detects
1.1 scanning electron microscopes detect
Preparation concentration is 2%, 3%, 4% gel sample, and gel thicknesses is 0.5cm.Gel in-40 DEG C of refrigerators more than pre-freeze 2h after, put into freezer dryer lyophilizing 24h.The gel sample of three concentration of lyophilizing is longitudinally cut, choose the morphosis by scanning electron microscopic observation gel surface after arbitrary facet metal spraying.
1.2 SIS thermosensitive hydrogel external degradation
3% the gel sample of preparation 1mL, puts into EP pipe, adds 1mLPBS or 0.05%I Collagenase Type solution, is positioned on 37 DEG C of shaking tables, in 0 day, 1 day, 5 days, 10 days, EP pipe is taken out in 20 days to centrifugal rear taking-up supernatant.After the lyophilizing of all EP pipes, weigh, the comparison degradation rate taking out during with 0 day.
The external slow release of 1.3 somatomedin: solution after collection SIS gel external degradation, ELISA detects its vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) Cumulative release amount.
2.SIS thermosensitive hydrogel Cyto-compatibility in vitro (taking commercially available NTx as contrast)
2.1 SIS thermosensitive hydrogel promote cell proliferation experiment
In 96 orifice plates, draw 3%SIS thermosensitive hydrogel and matched group NTx (COL) 50 μ L are laid in each hole, put into 37 DEG C of incubators and make its plastic.NIH 3T3 cell is become after cell suspension with HUVEC cell dissociation, regulate cell concentration, with the cell number in every hole 8000, cell is inoculated in to two kinds of gel surface in 96 orifice plates.Detected cell proliferation situation at the 1st day, 3 days, 5 days, 7 days by CCK-8 cell proliferation and toxicity detection kit.
2.2 LIVE/DEAD cell fluorescence dyeing
SIS thermosensitive hydrogel and NTx surface are with 10
5/ cm
2density inoculation HUVEC, cultivate after five days, add calcein (AMCalcein-AM) and propidium iodide (PI) fluorescence dye liquor, dye after 20min, under inverted fluorescence microscope, observe.
3.SIS thermosensitive hydrogel body is implanted into (inflammatory reaction evaluation) (taking commercially available NTx as contrast)
By subcutaneous to SIS thermosensitive hydrogel and NTx 1mL injection SD rat back, in 3 days, one week, two weeks, after surrounding, to draw materials together with skin of back, specimens paraffin embedding slices after paraformaldehyde is fixing, carries out HE dyeing.
4.SIS thermosensitive hydrogel Angiogensis (taking commercially available NTx as contrast)
4.1 SIS thermosensitive hydrogel promote HUVEC to form the experiment of lumen of vessels spline structure
In 24 orifice plates, drawing the each 90 μ L of 3%SIS thermosensitive hydrogel and NTx is laid in hole HUVEC with 1.5 × 10
5/ cm
2density inoculation, after 3 days, observe HUVEC and whether formed lumen of vessels spline structure.
4.2 SIS thermosensitive hydrogel promote rat aorta ring rudiment experiment
The SD rat chest aorta of one week is cut into the ring-type that about 1mm is thick, in 48 orifice plates, first spreads 50 μ LSIS gels, after plastic on glue face aortic annulus of parallel placement, the SIS gel that repaves one deck 50 μ L will encircle parcel.Add EBM-2 culture medium, cultivate after 10 days, observe vascular ring bud.Matched group NTx in like manner.
Short Angiogenesis experiment after injection in 4.3 SIS thermosensitive hydrogel bodies
SIS thermosensitive hydrogel and NTx 1mL are subcutaneously injected into rat back, within 3 days, drew materials together with skin of back after 7 days, specimens paraffin embedding slices after paraformaldehyde is fixing, by the mode labelling blood capillary (CD31) of immunohistochemical staining, observes the lower newborn blood capillary of material induction.
5 testing results:
5.1 scanning electron microscopes detect
After submucous layer of small intestine substrate is mixed with lysate, 37 DEG C is formation temperature gel (Fig. 1).Fig. 2 is followed successively by the scanning electron microscopic observation figure of 2%, 3%, 4% SIS thermosensitive hydrogel from left to right, and amplification is 500 ×.The aperture of gel is along with gel strength raises and reduces.
5.2 SIS thermosensitive hydrogel percent hydrolysiss detect
The results are shown in Figure 3.As shown in Figure 3,3%SIS thermosensitive hydrogel percent hydrolysis tended towards stability substantially after 10 days, 40% left and right of finally degrading in PBS.After 10 days, enzymolysis approaches 70%.
5.3 SIS gel growth factor slow release
The results are shown in Figure 4, without somatomedin initial stage outburst release phenomenon, extend with degradation time, growth factor release amount increases.
5.3 SIS thermosensitive hydrogel promote cell proliferation experiment
The results are shown in Figure 5. as shown in Figure 5, detect that by CCK-8 cell proliferation and toxicity detection kit HUVEC and two kinds of cells of NIH 3T3 all can, at SIS thermosensitive hydrogel and NTx (COL) surface growth, show two kinds of equal no cytotoxicities of gel.
5.4 LIVE/DEAD cell fluorescence dyeing
The results are shown in Figure 6.As shown in Figure 6, inoculate HUVEC1, after 3,5,7 days, do the dyeing of LIVE/DEAD cell fluorescence, SIS thermosensitive hydrogel and NTx (COL) surface has no a large amount of red dead cells, and cell compatibility is good.
5.5 SIS thermosensitive hydrogel bodies are implanted into (inflammatory reaction evaluation)
The results are shown in Figure 7,8.In Fig. 7, be respectively subcutaneous injection SIS thermosensitive hydrogel and NTx (COL) latter 3 days, 7 days, 2 weeks, after 4 weeks, draw materials, do HE coloration result.Fig. 8 is number of inflammatory cells order testing result.After visible injection SIS thermosensitive hydrogel, in the time of 3 days and 7 days, have slight inflammatory reaction, but inflammatory reaction disappears substantially during by 2 weeks.In whole process, having no obvious fibrous capsule forms.After 2 weeks, material is degraded in a large number, rarely seen a small amount of remnants 4 weeks time.Count and confirmed that inflammatory reaction is lighter by inflammatory cell, disappear very fast.In sum, SIS thermosensitive hydrogel has reduced immunogenicity, and biocompatibility is good.
5.6 SIS thermosensitive hydrogel promote HUVEC to form the experiment of lumen of vessels spline structure
With SIS temperature-sensitive hydrogel and NTx (COL) thermosensitive hydrogel surface seeding HUVEC 1 day, 3 days, after 5 days, use laser confocal microscope (40X) to observe, the results are shown in Figure 8.As shown in Figure 8, after inoculation, within the 1st day, be that visible HUVEC is arranged in network structure, but microvessel structure is not obvious.During by the 3rd day, HUVEC is further arranged in netted, and has blood capillary chamber spline structure to occur.During by the 5th day, lumen of vessels spline structure is obvious, keeps stable (Fig. 9 N).And the cell that is inoculated in NTx surface is only bred, have no and form blood vessel network structure (Fig. 9 M).SIS thermosensitive hydrogel can promote HUVEC to form blood capillary chamber spline structure, and short blood vessel occurs.
5.7 SIS thermosensitive hydrogel promote rat aorta ring rudiment experiment
The results are shown in Figure 10.As shown in Figure 10, when rat aorta ring is cultured to the 10th day, visible aortic annulus has blood capillary rudiment to occur around, and the vascular ring that NTx (COLL) is hatched has no significant change, and two groups of micro-vessel areas have significant difference (Figure 11).Visible, SIS thermosensitive hydrogel can promote rat aorta ring rudiment to occur.
Short Angiogenesis experiment after injection in 5.8 SIS thermosensitive hydrogel bodies
The results are shown in Figure 12, Figure 12 is rat back injection SIS thermosensitive hydrogel and NTx thermosensitive hydrogel (COL) embedded section of drawing materials after 3 days and 7 days, carries out immunohistochemical staining, and label is CD31, label vascular endotheliocyte.What in upper figure, brown color was circular is blood capillary newborn in material, and two kinds of gel injections can be induced the generation of new vessels after 3 days and 7 days.But blood capillary number and blood vessels caliber that the induction of SIS thermosensitive hydrogel generates are all obviously greater than NTx thermosensitive hydrogel (Figure 13), it promotes the ability of angiogenesis to be better than NTx as seen.
Claims (9)
1. the temperature sensing material of the promotion vascularization based on submucous layer of small intestine, is characterized in that: it is to prepare after submucous layer of small intestine gel matrix material, lysate are packed respectively, and wherein, submucous layer of small intestine gel matrix material obtains by the following method:
Get submucous layer of small intestine, through enzymolysis, acid-soluble, lyophilization, beats powder after dry or shreds into cotton-shapedly, and sterilizing, obtains aseptic powdery or cotton-shaped submucous layer of small intestine gel matrix material;
Lysate is phosphate buffer or the normal saline that contains NaOH.
2. temperature sensing material according to claim 1, is characterized in that: the mass volume ratio of submucous layer of small intestine gel matrix material and lysate is 2～4:100.
3. temperature sensing material according to claim 1 and 2, is characterized in that: in described lysate, the content of NaOH is to regulate submucous layer of small intestine gel matrix material pH=6.5～7.2 to be as the criterion.
4. temperature sensing material according to claim 1, is characterized in that: the concrete operations of preparing submucous layer of small intestine gel matrix material are as follows:
(1) get dry submucous layer of small intestine, pulverize, gained SIS powder is for subsequent use;
(2) in HCl solution, add pepsin, treat that pepsin dissolves completely, obtains Digestive system;
(3) get SIS powder and Digestive system, after mixing, be placed on shaking table and rock, after Digestive system becomes clarification, take out and carry out lyophilization, beat powder after dry or shred into cotton-shapedly, sterilizing, obtains aseptic powdery or cotton-shaped submucous layer of small intestine gel matrix material.
5. temperature sensing material according to claim 4, is characterized in that: in step (1), got the powder of 80 mesh sieves after pulverize at low temperature, and be SIS powder.
6. thermosensitive hydrogel according to claim 4, is characterized in that: in step (2), and HCl pH value of solution=2～4; In Digestive system, pepsin concn is 1mg/ml.
7. temperature sensing material according to claim 4, is characterized in that: in step (3), the mass volume ratio of SIS powder and Digestive system is 1:100～2:100.
8. according to the temperature sensing material described in claim 1 or 4, it is characterized in that: submucous layer of small intestine gel matrix material adopts irradiation or oxirane to carry out sterilizing, and lysate adopts filtration sterilization.
9. the preparation method of the temperature sensing material of the promotion vascularization based on submucous layer of small intestine described in claim 1, is characterized in that: it comprises following operating procedure:
(1) prepare submucous layer of small intestine gel matrix material:
A, get dry submucous layer of small intestine, pulverize, gained SIS powder is for subsequent use;
B, in HCl solution, add pepsin, treat that pepsin dissolves completely, obtains Digestive system;
C, get SIS powder and Digestive system, after mixing, be placed on shaking table and rock, after Digestive system becomes clarification, take out and carry out lyophilization, beat powder after dry or shred into cotton-shapedly, sterilizing, obtains aseptic powdery or cotton-shaped submucous layer of small intestine gel matrix material;
(2) after getting submucous layer of small intestine gel matrix material, lysate and packing respectively, obtain temperature sensing material.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410440422.0A CN104189009B (en) | 2014-09-01 | 2014-09-01 | Vascularization promoting submucous layer of small intestine temperature sensing material and preparation method thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410440422.0A CN104189009B (en) | 2014-09-01 | 2014-09-01 | Vascularization promoting submucous layer of small intestine temperature sensing material and preparation method thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN104189009A true CN104189009A (en) | 2014-12-10 |
| CN104189009B CN104189009B (en) | 2017-12-08 |
Family
ID=52074485
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201410440422.0A Active CN104189009B (en) | 2014-09-01 | 2014-09-01 | Vascularization promoting submucous layer of small intestine temperature sensing material and preparation method thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN104189009B (en) |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105126171A (en) * | 2015-08-27 | 2015-12-09 | 中国人民解放军第三军医大学 | Gel biological material having shape memory function and preparation method of gel biological material |
| CN105999373A (en) * | 2016-05-30 | 2016-10-12 | 四川大学华西医院 | Polyurethane/small intestinal submucosa (PU/SIS) dual-component injectable hydrogel |
| CN107308495A (en) * | 2017-07-06 | 2017-11-03 | 苏州期佰生物技术有限公司 | A kind of artificial blood vessel based on trees-Osima jacoti, Osima excavata and preparation method thereof |
| CN109513037A (en) * | 2018-11-14 | 2019-03-26 | 华中科技大学同济医学院附属协和医院 | A kind of submucous layer of small intestine Wound dressing of loaded mesoporous bio-vitric |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2008109407A2 (en) * | 2007-03-02 | 2008-09-12 | University Of Pittsburgh-Of The Commonwealth System Of Higher Education | Extracellular matrix-derived gels and related methods |
| CN101433735A (en) * | 2007-11-13 | 2009-05-20 | 北京大清生物技术有限公司 | Method for preparing SIS tissue repair material |
-
2014
- 2014-09-01 CN CN201410440422.0A patent/CN104189009B/en active Active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2008109407A2 (en) * | 2007-03-02 | 2008-09-12 | University Of Pittsburgh-Of The Commonwealth System Of Higher Education | Extracellular matrix-derived gels and related methods |
| CN101433735A (en) * | 2007-11-13 | 2009-05-20 | 北京大清生物技术有限公司 | Method for preparing SIS tissue repair material |
Non-Patent Citations (2)
| Title |
|---|
| KKOT NIM KANGA: "In vivo release of bovine serum albumin from an injectable small intestinal submucosa gel", 《INTERNATIONAL JOURNAL OF PHARMACEUTICS》 * |
| 罗静聪: "小肠黏膜下层的制备及其特性的研究进展", 《中国修复重建外科杂志》 * |
Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105126171A (en) * | 2015-08-27 | 2015-12-09 | 中国人民解放军第三军医大学 | Gel biological material having shape memory function and preparation method of gel biological material |
| CN105999373A (en) * | 2016-05-30 | 2016-10-12 | 四川大学华西医院 | Polyurethane/small intestinal submucosa (PU/SIS) dual-component injectable hydrogel |
| CN105999373B (en) * | 2016-05-30 | 2019-08-16 | 拓睿美（北京）医疗科技有限公司 | A kind of polyurethane/submucous layer of small intestine (PU/SIS) two-component injection aquagel |
| CN107308495A (en) * | 2017-07-06 | 2017-11-03 | 苏州期佰生物技术有限公司 | A kind of artificial blood vessel based on trees-Osima jacoti, Osima excavata and preparation method thereof |
| CN109513037A (en) * | 2018-11-14 | 2019-03-26 | 华中科技大学同济医学院附属协和医院 | A kind of submucous layer of small intestine Wound dressing of loaded mesoporous bio-vitric |
| CN109513037B (en) * | 2018-11-14 | 2021-09-17 | 华中科技大学同济医学院附属协和医院 | Mesoporous bioglass-loaded small intestine submucosa wound dressing |
Also Published As
| Publication number | Publication date |
|---|---|
| CN104189009B (en) | 2017-12-08 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN104582747B (en) | For manufacturing the method for the hydrogel fines with living cells and for manufacturing the compositions of tissue engineering bracket | |
| ES2668398T3 (en) | Compositions and procedures for tissue filling and regeneration | |
| CN102743796B (en) | Silk fibroin porous support made from polyvinyl alcohol, and preparation method and application thereof | |
| CN103877617B (en) | Two cross-linked hydrogel of injectable fibroin protein-alginate and preparation method thereof and using method | |
| CN102198292B (en) | Scaffolds of umbilical cord decellularized Wharton jelly for tissue engineering and preparation method thereof | |
| CN107224617B (en) | Hydrogel taking spleen extracellular matrix as raw material and preparation method thereof | |
| CN105963785B (en) | Acellular matrix material based on adipose-derived stem cell membrane and preparation method thereof | |
| CN102933705A (en) | Decellularized adipose tissue | |
| CN104353111B (en) | A kind of bioprosthetic material for abdominal-wall defect and preparation method thereof | |
| Kofidis et al. | Clinically established hemostatic scaffold (tissue fleece) as biomatrix in tissue-and organ-engineering research | |
| KR101889697B1 (en) | Muscle tissue regeneration using muscle fiber fragments | |
| CN102218160B (en) | Preparation and application of nerve tissue matrix derived tissue engineering scaffold material | |
| CN102973984B (en) | Preparation method and application of composite porous scaffold | |
| CN104055795B (en) | A kind of injectable implant and preparation method thereof | |
| CN104189009A (en) | Small intestinal submucosa-based vascularization promoting thermosensitive material and preparation method thereof | |
| CN112980001B (en) | Collagen composite hyaluronic acid gel, extracellular matrix bionic material and preparation method | |
| CN105251052A (en) | Cartilage extracellular matrix and silk fibroin composite orientation cartilage support and preparation method thereof | |
| Anton-Sales et al. | Bacterial nanocellulose and titania hybrids: cytocompatible and cryopreservable cell carriers | |
| CN104548200A (en) | Method of preparing highly branched polysaccharide-fibroin hydrogel bracket | |
| CN103599567B (en) | Temperature-sensitive composite as well as preparation method and application thereof | |
| CN107158463A (en) | Preparation method and purposes for the pancreatic parenchymal hydrogel of repairing pancreas tissue | |
| CN104056304A (en) | Growth-factor-chitosan-microsphere loaded DBM support joint cartilage repairing material | |
| CN105126171A (en) | Gel biological material having shape memory function and preparation method of gel biological material | |
| CN107693844A (en) | A kind of composition gels and application | |
| Li et al. | L-polylactic acid porous microspheres enhance the mechanical properties and in vivo stability of degummed silk/silk fibroin/gelatin scaffold |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant | ||
| CB03 | Change of inventor or designer information | ||
| CB03 | Change of inventor or designer information |
Inventor after: Luo Jingcong Inventor after: Wang Wei Inventor after: Ding Wei Inventor before: Luo Jingcong Inventor before: Wang Wei |